We propose to examine whether alterations in the lipid fluidity of the adipocyte plasma membrane occur in 'insulin-resistant', hyperglycemic states. Fat cell ghosts and purified plasma membrane fractions will be labeled with fatty acid and phospholipid spin probes, I(m,n). Electron spin resonance (ESR) spectra of the I(m,n)-labeled membrane provide information on the fluidity of the membrane lipid environment of the probe. We will study the structures of I(m,n)-labeled adipocyte membranes prepared from insulin-deficient, streptozotocin diabetic rats and also from hyperinsulinemic, obese hyperglycemic (ob/bo) mice. Basel and insulin-stimulated glucose uptake will be assessed in the intact isolated cells. The uptake of glucose will be measured through the use of 2-deoxy-(1-14C) glucose and D-(U-14C) glucose and also by determining the removal of unlabeled hexose from the incubation medium. Any alterations in the membrane fluidity might be related to the hyperglycemia and the resistance to insulin action of adipocytes from the diabetic (hyperglycemic) animals. In addition, membrane structural/functional aberrations resulting from chronic insulin deficiency might be reversing by treating the streptozotocin diabetic animals with insulin. Studies will be conducted to determine if the lipid fluidity is involved in the stimulation of adipocyte glucose uptake by insulin. A variety of lipid perturbing agents such as local anesthetics, both neutral and charged, the cations Ca 2 ion, Mg 2 ion, and La 3 ion, cholesterol, the cholesterol-complexing drugs filipin and digitonin, and temperature alterations will be tested for their effects on the membrane lipid fluidity. In parallel functional studies, the effects of these perturbants on the basal and insulin-stimulated uptake of glucose by isolated adipocytes will be determined. The possibility that insulin mediates direct effects on the fluidity of the adipocyte bilayer will be studied by comparing I(m,n)-labeled membranes prepared from insulin-treated adipocytes. These studies may help define the involvement of the membrane lipid structure in the mechanism of action of insulin and may determine whether the lipid fluidity modulates fat cell functions.